Apoptosis and necrosis represent two different mechanisms of cell death. Apoptosis is a highly regulated process involving the caspase family of cysteine proteases, and characterized by cellular shrinkage, chromatin condensation, and DNA degradation. In contrast, necrosis is associated with cellular and organelle swelling and plasma membrane rupture with ensuing release of intracellular contents and secondary inflammation (Kroemer et al., Cell Death Differ., 16:3-11 (2009)). Necrosis has been considered a passive, unregulated form of cell death; however, recent evidence indicates that some necrosis can be induced by regulated signal transduction pathways such as those mediated by receptor interacting protein kinases (RIPKs) especially in conditions where caspases are inhibited or cannot be activated efficiently (Golstein, P. et al., Trends Biochem. Sci., 32:37-43 (2007); Festjens et al., Biochim. Biophys. Acta, 1757:1371-1387 (2006)). Stimulation of the Fas and TNFR family of death domain receptors (DRs) is known to mediate apoptosis in most cell types through the activation of the extrinsic caspase pathway. In addition, in certain cells deficient for caspase-8 or treated with pan-caspase inhibitor Z-VAD, stimulation of death domain receptors (DR) causes a receptor interacting protein kinase 1 (RIPK1) dependent programmed necrotic cell death instead of apoptosis (Holler et al., Nat. Immunol., 1:489-495 (2000); Degterev et al., Nat. Chem. Biol., 4:313-321 (2008)). This novel mechanism of cell death is termed “programmed necrosis” or “necroptosis” (Degterev et al., Nat. Chem. Biol., 1:112-119 (2005)).
Necroptosis can be triggered upon activation of TNF receptors or Toll-like receptors in response to genotoxic stress and during virus infection and has been shown to be RIPK1 and RIPK3 dependent. Studies reveal that the expression of RIPK3 and the RIPK1-RIPK3 binding through the RIP homotypic interaction motif (RHIM) is a prerequisite for RIPK1 activation, leading to reactive oxygen species (ROS) production and necrotic cell death (He et al., Cell, 137:1100-1111 (2009); Cho et al., Cell, 137:1112-1123 (2009); Zhang et al., Science, 325:332-336 (2009)).
Dysregulation of RIPK3-dependent signaling has been linked to inflammatory diseases such as macrophage necrosis in atherosclerosis development, virus-induced inflammation, systemic inflammatory response syndrome and ethanol-induced liver injury, neurodegeneration such as detachment of the retina, ischemia, and Gaucher's disease (Trichonas et al., Proc. Natl. Acad. Sci., 107:21695-21700 (2010); Lin et al., Cell Rep., 3:200-210 (2013); Cho et al., Cell, 137:1112-1123 (2009); Duprez et al., Immunity, 35:908-918 (2011); Roychowdhury et al., Hepatology, 57:1773-1783 (2013); Vandenabeele et al., Nature, 10:700-714 (2010); Vandenabeele et al., Sci. Signalling, 3:1-8 (2010); Zhang et al., Cell. Mol. Immunol., 7:243-249 (2010); Moriwaki et al., Genes Dev., 27:1640-1649 (2013); Vitner et al., Nat. Med., 20:204-208 (2014)).
A potent, selective, small molecule inhibitor of RIPK3 activity would block RIPK3-dependent pro-inflammatory signaling and thereby provide a therapeutic benefit in inflammatory diseases characterized by increased and/or dysregulated RIPK3 kinase activity.